The long-term research goal of the proposed project is to understand the pathophysiologic mechanisms of progressive connective tissue breakdown widely experienced in gingivitis and periodontitis. Type XII collagen is a homotrimer made up of alpha1 (XII) chains and found to be present in the periodontal ligament, and in tendons and ligaments, tissues which predominantly contain type I collagen. This molecule belongs to a recently found group of collagens that represent minor constituents of the extracellular matrix and are referred to collectively as FACITs or Fibril Associated Collagens with Interrupted Triple-Helices. The tissue- specific expression of FACITs and the characteristic heterogeneity in their amino-terminal domains strongly suggest that FACITs may play a significant role in organizing the three-dimensional spatial pattern of collagen fibrils in different tissues. As described in Preliminary Studies we have shown that the expression of type XII collagen is significantly associated with the rat PDL and dentogingival fiber regions during the maturing stage. In the healthy periodontium, the dentogingival fibers are arranged in neat, parallel bundles. During the initial stages of inflammation each individual fiber maintains its integrity, but the parallel arrangement is disrupted and the fibers appear dissociated from one another. In this application, we proposed a hypothesis that type XII collagen may be degraded first in gingivitis and periodontitis, leading to collagen fibril disintegration. This molecule may therefore play a significant role in the pathophysiology of progressive periodontal tissue breakdown. As the first step to test our hypothesis, the expression of type XII collagen mRNA will be studied in a well established periodontitis model in the rat. Next, the primary structure of rat XII collagen will be analyzed by molecular cloning and sequencing of rat alpha1 (XII) cDNAs using a primer extension method. The DNA sequence of alpha1(XII) collagen cDNAs will provide deduced peptide sequences based on which several synthetic oligo-peptides will be prepared. The synthetic oligo- peptide will be further used to generate affinity purified polyclonal antibodies against rat type XII collagen. Finally, studies of immunoblotting and immunohistochemistry will be carried out in various stages of gingivitis and periodontitis using the generated anti-rat type XII collagen antibodies. The findings from this project will provide the basis for better understanding of the specific pathophysiologic mechanisms of periodontal disease at the molecular level. Furthermore, both cDNAs and antibodies recognizing rat type XII collagen mRNA/DNA and protein, respectively, can potentially serve as significant tools to investigate the aspects of aging, response to infection, effect of systemic diseases, and genetic disturbances on the periodontium. These studies will serve as an important background for human disease.